Most eukaryotic cells contain mitochondria, the organelles required to produce cellular energy. In addition to the nuclear genome, mitochondria contain DNA and the maintenance of the mitochondrial genome is essential to the normal function of the cell. Mutations in mitochondrial DNA are associated with several human neuromuscular diseases, with age-related illnesses, such as diabetes mellitus, and with the aging process itself. The long-term goal of this project is to identify factors that are involved in the replication and repair of mitochondrial DNA using Saccharomyces cerevisiae as a model system. Mutants will be generated that increase the instability of mitochondrial microsatellite sequences. Microsatellites are regions in which a single base, or a small number of bases is repeated multiple times. Such simple repetitive sequences are abundant in eukaryotic genomes and are sensitive indicators of genomic stability because they alter at a much higher rate than non-repetitive sequences. Another approach is to determine the effect of mutations in genes whose products are known, or suspected, to be involved in DNA replication and repair in the mitochondrial compartment. In addition, studies will address the properties of the DNA sequences that affect their stable maintenance in the mitochondrial genome. The study of genes, whose products affect the stability of microsatellite sequences in the nucleus of yeast, led to the identification of the human homologs of some of these genes. Mutations in these human genes have been found to be involved in the predisposition to cancer. The experiments in this proposal are aimed at understanding the processes of mutagenesis and repair of mitochondrial DNA in yeast, however, these studies may lead to insights into similar processes in human mitochondrial DNA.